Rheumatoid arthritis (RA) is a chronic inflammatory disease, generally regarded as an autoimmune disorder, that affects approximately 1% of the adult population. It is characterized primarily by inflammation of the peripheral joints, in many cases ultimately leading to destruction of these joints. However, RA is a systemic disease, as especially long-standing (and severe) cases also develop extra-articular manifestations of symptoms. As the structural damage is progressive and largely irreversible, it is important to diagnose RA as early as possible to be able to start an adequate treatment. This holds especially true for patients at risk of or having severe RA, which is characterized, for example, by increased joint destruction (as measured by a higher radiological progression rate).
RA is caused by Chronic Inflammation of the Synovium that does not heal (Firestein, Nature, 423:356-361 (2003)). The synovium is a thin layer of tissue composed of 3 to 4 layer of cells that form a membrane encapsulating the joint fluid in a synovial joint (Iwanaga et al., Arch. Histol. Cytol., 63:17-31 (2000)). Chronic inflammation of the synovium leads to unchecked proliferation of the synovial tissue, which is composed mainly of two types of cells, the A cells which are macrophage-like and the B cells which are fibroblast-like (Iwanaga et al., supra). The outgrowth of the proliferating synovium into the joint cavity causes swelling of the joint and the formation of a destructive piece of hanging tissue called the “Pannus” (Sanchez-Pernaute et al., Rheumatology, 42:19-25 (2003)). It is the Pannus which acts like a “warhead” of a missile that does most of the damage to the articular cartilage by the generation and secretion of the matrix metalloproteinases (MMPs) that break up the cartilage matrix proteins in the joint (Pap et al., Arthritis Res., 2:361-367 (2000); Konttinen et al., Matrix Biology, 17:585-601 (1998)). Once the cartilage starts to erode, the disease becomes very serious because joint swelling and pain starts to develop which is the hallmark of RA (Firestein, Nature, 423:356-361 (2003)).
However, several fundamental questions on the etiology of RA remain. It is still unknown what mechanism initiates the inflammation in the synovium, and similarly, what mechanism sustains the chronic inflammation in the synovium. What is known, is that chronic inflammation in any bodily tissue has to be driven and sustained by the continuous presence of a foreign antigen (or antigens) in the inflamed tissue. Unfortunately, no animal models of arthritis can truly mimic the chronic inflammatory condition of RA because the so-called “collagen-induced arthritis”, “albumen-induced arthritis” and “bacterial cell wall-induced arthritis” animal models of arthritis (Brahn Clin. Orthop. Relat. Res., 265:42-53 (1991)) are all acute animal models, as the arthritic disease is induced by immunizing the animals with a subcutaneous injection of the antigen mixed with Freund's complete adjuvant and then followed later by an injection of the antigen into the synovial joint cavity in one of the knees of the animal while the non-injected knee serves as the control. After the injection, arthritic disease will develop in the injected knee joint within a week while the non-injected knee joint is disease-free. However, appearance of the disease is only transient because the diseased joint will eventually heal itself and the animal recovers spontaneously. This phenomenon occurs because the antigen that causes the disease is no longer present in the diseased joint to sustain the disease. As such, in all of the animal models of arthritis, the antigen that induces the disease is delivered exogenously by manual injection into the joint cavity. Thus, in order to properly diagnose and treat RA it is of paramount importance to determine what mechanism(s) are responsible for generating and sustaining a continuous presence of one or more foreign antigens, as well as the identity of the foreign antigen(s) underlying the disease.
Due to the tremendous research efforts executed by numerous laboratories all over the world in the past 15 years, researchers believe that the foreign antigen (or antigens) responsible for inducing and sustaining RA in RA-prompt patients are the citrulline-containing peptides derived from citrullination of the endogenous cellular proteins by the intracellular enzymes, peptidyl-arginine deiminases (PADs) (Schellekens et al., J. Clin. Invest., 101:273-281 (1998); Girbal-Neuhauser et al., J. Immunol., 162:585-594 (1999)). PADs convert an arginine residue within a peptide sequence to a citrulline residue and this reaction only occurs in the presence of >10−4M Ca2+ concentration. There are five known members of PADs (I, II, III, IV and VI) present inside the cells, and PADs II and IV are the ones found in the synovial cells (Foulquier et al., Arthritis & Rheumatism, 56:3541-3553 (2007)). Once a PAD is released outside of the cell, it can citrullinate other extracellular proteins but the enzymatic activity also disappears rapidly. Therefore, to further explore the cause of RA, one has to determine how the citrullinated peptides are being generated in the synovium and what sustains the continuous generation of those citrullinated peptides.
A correct diagnosis of RA is often difficult because the symptoms develop insidiously, or may resemble those of other diseases (e.g., osteoarthritis, arthritis due to infection or gout, etc.). Traditionally, RA is diagnosed using the revised American College of Rheumatology (ACR) classification criteria. The ACR proposes seven classification criteria which indicate a poor prognosis:                1. Morning stiffness of the joints lasting more than one hour;        2. Arthritis of three or more joints;        3. Inflammation of at least three joint areas at the same time;        4. Hand joints or finger joints are likewise affected;        5. Bilateral tenderness of metacarpophalangeal joints to pressure;        6. Erosions on radiographs;        7. Detection of rheumatoid factors, anti-perinuclear factor (APF), and anti-keratin antibodies (AKA).However, diagnosing RA according to this procedure is labor-intensive and a significant amount of time passes before a definite diagnosis is made.        
Autoantibodies to the “anti-perinuclear factor” (APF) were first described by Nienhuis et al. in patients having rheumatoid arthritis (Nienhuis et al., Ann. Rheum. Dis., 23:302-305 (1964)). These APF antibodies react with the keratohyaline scattered around the perinuclear region of human buccal epithelial cells. Owning to the subjective and labor-intensive immunofluorescence technique employed, an APF antibody test has never been put into wide use for RA diagnosis. Later, Young et al. reported that RA patient sera reacted to the keratinous epithelium of the stratum corneum on rat esophagus tissue sections and designated these RA-specific antibodies as anti-keratin antibodies (AKA) (Young et al., B.M.J., 2:97-99 (1979)). In 1993, Simon et al. found that a majority of the RA patient sera recognized a 40 kDa protein from human skin tissue (Simon et al., J. Clin. Invest., 92:1387-93 (1993)). They further demonstrated that this protein identified as filaggrin was the target antigen of AKA and went on to show that AKA and APF antibodies are the same RA-specific antibodies (Sebbag et al., J. Clin. Invest., 95:2672-2679 (1995)). For this reason, the APF autoantibodies are even today referred to as antikeratin antibodies (AKAs) (Vincent et al., J. of Autoimmunity, 4:493-505 (1991); Paimela et al., Ann. Rheumat. Dis., 51:743-746 (1992)). The 40 kDa filaggrin protein aggregates cytokeratin filaments and assists in forming the intracellular fiber matrix of the keratinous cells (Simon et al., J. Clin. Invest., 92:1387-93 (1993)). However, filaggrin is not present in the synovial joint tissue of RA patients. Furthermore, anti-filaggrin antibodies are found in the serum of only about 40% of RA patients.
As such, there is a need in the art for the identification and design of novel peptides that find utility in detecting antibodies associated with rheumatic diseases, e.g., antibodies associated with RA, which peptides make possible a sensitive and specific diagnosis, classification, and/or prognosis of rheumatic diseases such as RA. The present invention satisfies this need and provides related advantages as well.